摘要
目的分析结直肠癌患者与健康人的粪便微生物群落,以及结直肠癌组织与癌旁组织菌群差异,期望发现肠道内菌群差异与结直肠癌的相关性。方法选取2021年9月至2022年10月包头市中心医院收治的结直肠癌患者和健康者,共收集60例样本,其中收集结直肠癌患者粪便11份,记为A组。采集结直肠癌组织20份,对应的癌旁组织20份,分别记为B组、C组。同时收集健康人粪便9份,记为D组。送测序公司进行菌群测序,分析A组与D组、B组与C组之间菌群的组成及多样性情况。结果共获得4335646条高质量扩增序列,按97%的序列相似度进行归并,与Greengenes数据库进行物种注释对比,生成特征性序列OTU和丰度数据表格。多样性稀释曲线趋于平缓,表明本次测序覆盖了绝大多数物种。在门水平上,优势菌在A、B、C、D组中大致一致,其中壁厚菌门(Firmicutes)、拟杆菌门(Bacteroidetes)、变形菌门(Proteobacteria)占据了肠道菌群的90%以上,A组与D组、B组与C组间未见相对丰度有差异的细菌。在属水平上,Lachnobacterium在D组相对丰度高于A组,且差异有统计学意义(Z=-2.309,P=0.027),瘤胃球菌(Ruminococcus)在A组相对丰度高于D组,且差异有统计学意义(Z=-2.309,P=0.027)。Chao1指数反映物种丰富度,Shannon指数反映物种多样性。A组的Chao1指数低于D组,且差异有统计学意义(Z=-3.001,P=0.003)。Chao1指数在B组与C组间差异无统计学意义。对于Shannon指数,A组与D组、B组与C组间差异均无统计学意义。对于菌群beta多样性,A组、D组之间菌群的beta多样性上存在着显著结构差异,且差异具有统计学意义(F=2.684,P=0.004),但B组、C组之间beta多样性差异无统计学意义。对于菌群差异物种筛选,A、D两组间相比,结直肠癌组(A组)出现6个差异物种,分别为瘤胃球菌属(g_Ruminococcus)、f_Haliangiaceae、f_[Tissierellaceae]、梭杆菌门(p_Fusobacteria)、梭杆菌目(o_Fusobacteriales)、梭杆菌纲(c_Fusobacteriia)。在健康对照组(D组)中,鉴定出14个差异物种,分别为厚壁菌门(p_Firmicutes)、梭菌目(o_Clostridiales)、梭状芽胞杆菌纲(c_Clostridia)、g_Lachnobacterium、疣微菌科-瘤胃球菌属(f_Ruminococcaceae_g_Ruminococcus)、毛螺菌属(g_Lachnospira)、f_Christensenellaceae、交替单胞菌目(o_Alteromonadales)、柔膜菌纲(c_Mollicutes)、软壁菌门(p_Tenericutes)、o_RF39、嗜热油菌纲(c_Thermoleophilia)、副球菌属(g_Paracoccus)、诺卡菌科(f_Nocardiaceae)。对于组织样本,经过LEfSe分析后,未见结直肠癌组织与癌旁组织间存在差异物种。结论在门、属水平上优势物种和相对丰度存在差异,其中梭杆菌可能是参与结直肠癌的发生菌群。结直肠癌患者菌群的丰富度显著降低,患者与健康者相比,存在菌群群落的结构分离,同时结直肠癌患者的产短链脂肪酸的细菌相对丰度显著减少,以及机会致病菌的增多,提示结直肠癌患者存在一定的菌群失衡现象。结直肠癌组织与癌旁组织的菌群相对丰度、多样性未见明显的改变,菌群的结构也无明显分离,提示菌群在癌与癌旁组织之间可能不存在差异。
Objective To analyze the fecal microbial communities of colorectal cancer(CC)patients vs healthy people,and the differences in microflora between CC tissues and adjacent tissues,hoping to find the correlation between intestinal microflora differences and CC.Methods Patients admitted to the hospital from September 2021 to October 2022 were selected.A total of 60 samples were collected,among which 11 stool samples were collected from colorectal tumor patients and recorded as group A.Twenty samples of CC tumor tissue and 20 samples of corresponding paracancer tissue were collected,which were respectively recorded as group B and group C.Meanwhile,9 fecal samples from healthy individuals were collected and recorded as group D.Microflora sequencing was performed to analyze the composition and diversity of microflora as well as the differences among group A and group D,group B and group C.Results A total of 4,335,646 high quality amplified sequences were obtained,which were merged according to 97%sequence similarity and compared with the Greengenes database for species annotation to generate the characteristic sequence OTUs and abundance data tables.The dilution curve of diversity tended to flatten out,indicating that the sequencing covered most species.At phylum level,dominant bacteria were roughly consistent in groups A,B,C and D,among which Firmicutes,Bacteroidetes and Proteobacteria accounted for more than 90%.No bacteria with different relative abundance were found between group A and group D,and between group B and group C.On the genus level,the relative abundance of Lachnobacterium in group D was significantly higher than that in group A(Z=−2.309,P=0.027),and that of Ruminococcus in group A was significantly higher than that in group D(Z=−2.309,P=0.027).Chao1 index in group A was significantly lower than that in group D(Z=−3.001,P=0.003).Chao1 index showed no statistical significance between group B and group C.For Shannon index,there was no significant statistical difference between group A and D,and between group B and C.There was a significant structural difference in the beta diversity between groups A and D(F=2.684,P=0.004),but not in the beta diversity between groups B and C.As for the screening of bacterial flora differential species,there were 6 different species in CC group(group A)compared with group A and D,including g_Ruminococcus,f_Haliangiaceae,f_[Tissierellaceae],p_Fusobacteria,o_Fusobacteriales and c_Fusobacteriia.In the healthy control group(group D),14 distinct species were identified,including p_Firmicutes,o_Clostridiales,c_Clostridia,g_Lachnobacterium,f_Ruminococcaceae_g_Ruminococcus,g_Lachnospira,f_Christensenellaceae,o_Alteromonadales,c_Mollicutes,p_Tenericutes,o_RF39,c_Thermoleophilia,g_Paracoccus,f_Nocardiaceae.For tissue samples,no differential species was found between CC tissues and paracancer tissues after LEfSe analysis.Conclusion There are differences in the dominant species and relative abundance at phylum and genus levels,among which Clostridium may be involved in the occurrence of CC.The flora richness of CC patients decreased significantly.Compared with healthy individuals,there was structural separation of flora community,and the relative abundance of short-chain fatty acids producing bacteria in CC patients decreased significantly,while opportunistic pathogens increased,suggesting that there is a certain imbalance of flora in CC patients.There are no significant changes in the relative abundance and diversity of flora in CC tissue and adjacent tissues,and no obvious separation of flora structure,suggesting that there may be no difference in the flora between cancer and adjacent tissues.
作者
梁志
董振宇
乌日嘎
李鹏达
杜明月
郭静
侯润杰
王继军
LIANG Zhi;DONG Zhenyu;WU Riga;LI Pengda;DU Mingyue;GUO Jing;HOU Runjie;WANG Jijun(Baotou Central Hospital,Baotou,Inner Mongolia 014000,China;不详)
出处
《中国微生态学杂志》
CAS
CSCD
北大核心
2024年第8期900-906,共7页
Chinese Journal of Microecology
基金
包头市青年人才创新项目([2019]116号)。
